The role of oleic acid in the process of induced destruction of Escherichia coli M-17

The influence of oleic acid on the process of the destruction of E. coli, induced by the transfer of the bacteria from the growth medium into solution without nutrient substances and their treatment with oleic acid at a temperature of 45 degrees C, was studied. Oleic acid at a concentration of 200 nmol/ml accelerated the destruction of E. coli in comparison with the control organisms, but did not lead to the significant increase of the final degree of the destruction of biomass. The dynamics of the inclusion of labeled oleic acid into E. coli cell fractions after the induction of autolysis was analyzed. The radioactive label was most actively included into the lipid (chloroform) fraction of bacterial biopolymers. Simultaneously with the destruction of some cells in the population the inclusion of labeled oleic acid into intact cells was found to occur.     

Isolation and identification of growth stimulators of Escherichia coli M-17

The stimulating activity of E. coli M-17 culture fluid (CF) is determined by the action of low-molecular exometabolites, readily soluble in water. The high-molecular fraction was removed from CF by ultrafiltration, the growth stimulators were adsorbed on anion exchange resin and eluated at pH 3.1. In further purification HPLC and chromatography on TSK gel HW-40 were used. The identification of compounds was carried out by the methods of thin-layer chromatography, amino acid analysis and 1H-NMR spectroscopy. Glutamic acid (glutamine) and succinic acid proved to be the most active growth autostimulators. The data of biological testing made it possible to believe that CF also contained less active stimulators and/or synergic substances which had no their own activity, but stimulated growth, acting jointly with other compounds. In view of the definite specificity of action observed in initial CF, some differences in the spectrum of growth stimulators of other E. coli strains may be supposed.     

Changes in cell diameter during the division cycle of Escherichia coli

Extensive measurements of steady-state populations of several Escherichia coli strains have consistently indicated that cell diameter decreases with increasing cell length. This was observed both after electron microscopy of air-dried cells and after phase-contrast microscopy of living cells. The analysis was made by considering separately the unconstricted cells and three classes (slight, medium, and deep) of constricted cells in the population. During slow growth, cells with the average newborn length were up to 8% thicker than unconstricted cells twice as long. This decrease in diameter is less at higher growth rates. Despite the small changes and the large variation of the diameter in any particular length class, significant negative correlations between diameter and length were obtained. Cell diameter increases again at the end of the cell cycle as indicated by an increase of average diameter in the three consecutive classes of constriction.     

Changes in the composition of membrane phospholipids during the cell cycle of Escherichia coli

Phospholipid concentrations have been estimated throughout the successive cell cycle in synchronously growing culture of E. coli B/r. Total phospholipid phosphorus was shown to be doubled in the period of time between two cell divisions, whereas during the division itself it did not change. Phosphatidylethanolamine (PE) and phosphatidylglycerol (PG) exhibit a stepwise increase during the cell cycle. It should be noted that the phase of accumulation of these lipids could shift depending on the duration of the cell cycle. The fall in level of PE was followed by a short-term increase (5-10 min). At the same time the level of cardiolipin was observed to be significantly increased.     

Changes in cell dimensions during amino acid starvation of Escherichia coli.

Electron microscopic analysis was used to study cells of Escherichia coli B and K-12 during and after amino acid starvation. The results confirmed our previous conclusion that cell division and initiation of DNA replication occur at a smaller cell volume after amino acid starvation. Although during short starvation periods, the number of constricting cells decreased due to residual division, it appears that during prolonged starvation, cells of E. coli B and K-12 were capable of initiating new constrictions. During amino acid starvation, cell diameter decreased significantly. The decrease was reversed only after two generation times after the resumption of protein synthesis and was larger in magnitude than that previously observed before division (F. J. Trueba and C. L. Woldringh, J. Bacteriol. 142:869-878, 1980). This decrease in cell diameter correlates with synchronization of cell division which has been shown to occur after amino acid starvation.     

Dynamics and functions of exometabolites in the process of the growth of Escherichia coli M-17 periodical culture

The concentrations of metabolites altered immediately after culture inoculation into fresh medium and depended on the their dilution (in small dilutions metabolites were released into the medium, and, on the contrary, in great dilutions they were adsorbed). Acetate was the main exometabolite. The growth of culture started only when the acetate concentration reached the threshold value. At the stage of growth deceleration an intensive release of acetate and other exometabolites occurred. The process had the explosive character of chemical chain reaction. It could be initiated by high local concentrations of metabolites around bacteria, most sensitive to changes in growth conditions. At a high concentration of such cells the signal could be transmitted from one bacteria to another, otherwise the reaction chain was quickly broken and no mass release of metabolites occurred. At the stationary phase succinate, valine and lactate were consumed by bacteria with lactate concentration dropping practically to 0. At the same time acetate and citrate concentrations oscillated about some constant value. The release of growth inhibitors, formiate and alanine, continued to the end of cultivation, which was, seemingly, one of the factors of growth cessation. The specific features of the dymamics of metabolites were indicative of the fact that they had their individual regulatory functions. The problem of the self-synchronization of the bacterial culture is discussed.     

Changes in buoyant density and cell size of Escherichia coli in response to osmotic shocks.

The buoyant density of Escherichia coli was shown to be related to the osmolarity of the growth medium. This was true whether the osmolarity was adjusted with either NaCl or sucrose. When cells were grown at one osmolarity and shocked to another osmolarity, their buoyant density adjusted to nearly suit the new osmolarity. When cells were subjected to hyperosmotic shock, they became denser than expected. When cells were subjected to hypoosmotic shock they occasionally undershot the new projected density, but the undershoot was not as dramatic as the overshoot seen with hyperosmotic shocks. Shrinkage and swelling of the cells in response to osmotic shocks could account for the change in their buoyant density. The changes in cell size after osmotic shocks were measured by two independent methods. The first method measured cell size with a Coulter Counter, and the second method measured cell size by stereologic analysis of Nomarski light micrographs. Both methods gave qualitatively similar results and showed the cells to be flexible. The maximum swelling recorded was 23% of the original cell volume, while the maximum shrinkage observed was 33%.     

Variations in cell size and buoyant density of Escherichia coli K12 during glycogen accumulation

The effect of glycogen accumulation on buoyant density and volume of Escherichia coli K12 was studied. A procedure consisting of three linear equations is presented. This requires measurement only of three parameters: cell buoyant density, cell volume and specific content of the polymer. Experimental values are then used to calculate intercepts and slopes of the equations by linear regression. From the estimated values of such parameters the in vivo values of several variables of interest can be calculated. These include in vivo density and volume of the glycogen inclusion, as well as density and volume of the structural material in the cell. The results are consistent with the glycogen inclusions being hydrated.     

Heterogeneity of Escherichia coli population during induced autolysis

Escherichia coli M-17 autolysis was induced by eliminating nutrition sources from the growth medium and exerting a shock with EDTA. The overall cell number, the optical density of the cell suspension, the number of colony-forming units (CFU), and [3H]uracil incorporation into the cells were analysed in the course of autolysis. The number of CFU was found to drop down faster than the overall cell number in the process of autolysis. The population of E. coli was shown to be heterogeneous in its sensitivity to the induction of autolysis, and some nonlysed cells were still metabolically active. When the rate of autolysis was highest in some cells of the population, the labeled precursor was found to be incorporated into the TCA-soluble and TCA-insoluble fractions of nonlysed cells. The overall cell number, the optical density of the cell suspension, and the number of CFU increased 96 h after the induction of autolysis. The authors discuss what is the role played by the heterogeneity of an E. coli population in its adaptation to EDTA-induced autolysis.     

Coupling the initiation of chromosome replication to cell size in Escherichia coli

Bacterial cells change size dramatically with change in growth rate, but the ratio between cell volume and the number of copies of the origin of chromosome replication (oriC) is roughly constant at the time of initiation of DNA replication at almost all growth rates. Recent research on the inactivation of initiator protein (DnaA) and depletion of DnaA pools by the high-affinity DnaA-binding locus datA allows us to propose a simple model to explain the long-standing question of how Escherichia coli couples DNA replication to cell size.     

Changes in prostaglandin production and ovarian function in gilts during endometritis induced by Escherichia coli infection

The aim of this study was to determine the prostaglandins (PGs) production and ovarian function in gilts after intrauterine infusions of 10(6) and 10(9) colony-forming units (cfu)/ml of Escherichia coli (E. coli). In Experiments 1 and 2, 30 ml of saline or 30 ml of E. coli suspension containing 10(6) or 10(9)cfu/ml, were infused once into each uterine horn in three groups of gilts on day 3 of the estrous cycle, respectively. In Experiment 1, 17 days after treatment it was revealed that inoculation of E. coli 10(9)cfu/ml induced severe acute or subacute endometritis while 10(6)cfu of E. coli evoked moderate acute endometritis or resulted in no inflammatory changes. In the gilts receiving 10(9)cfu/ml of E. coli, the concentration of 13,14-dihydro-15-keto-PGF(2)alpha in blood from the jugular vein was elevated (P<0.05-0.001) compared to concentration in the gilts inoculated with 10(6)cfu on days 8-17 after treatment. Both the E. coli-treated groups had a lower (P<0.05, P<0.01) progesterone plasma level from days 10 to 14 after administration than the control group. On day 17 of the study, infusion of E. coli 10(9)cfu/ml, in comparison to 10(6)cfu, resulted in the greater (P<0.001) content of PGE(2) in the myometrium. The content of both PGs in the endometrium as well as PGF(2alpha) in the myometrium of gilts-treated with 10(9)cfu/ml of E. coli was lower (P<0.001) than in gilts-treated with 10(6)cfu of bacteria. Newly formed corpora lutea were found in the gilts infused with 10(6), but not those infused with 10(9)cfu/ml of E. coli on day 17 after infusion. On day 8 of the study (Experiment 2), the blood from utero-ovarian vein of the gilts-treated with 10(9)cfu/ml of bacteria had a higher (P<0.05) PGF(2alpha) level and lower (P<0.001) PGE(2) level than following infusion of E. coli 10(6)cfu/ml. Also on day 8 of the study, the content of PGE(2) in the endometrium, both the PGs in the myometrium as well as cyclooxygenase-2 in the endometrium and myometrium was greater (P<0.01, P<0.001) after applying 10(9)cfu/ml than 10(6)cfu/ml of E. coli. These results indicate that intrauterine infusions of 10(6) or 10(9)cfu/ml of E. coli lead to the development of inflammatory states of different intensities which is connected with different PGF(2alpha) and PGE(2) production and function of ovaries.     

Changes in cell size and shape in thymine-requiring Escherichia coli associated with growth in low concentrations of thymine.

Thymine auxotrophs of three unrelated strains of Escherichia coli (K-12, B/r, and 15) were grown in media containing various concentrations of thymine. During steady-state growth conditions, cell volume increased as thymine concentration decreased but, in contrast to previous reports, this change was due to an increase in cell length without change in cell diameter.     

Comparative study of effects of Escherichia coli M-17 exometabolites and fructooligosaccharides on the growth and antagonistic activity of Lactobacilli

Facts concerning the evaluation of the influence of E. coli M17 exometabolites and fructooligosaccharides (FOS) on the growth and antagonistic activity of lactobacilli are presented. As revealed by these facts, preparation "Aktoflor" accelerates the growth of lactobacillary cultures, increases the final yield of biomass and antagonistic activity. E. coli M17 exometabolites contained in "Aktoflor" have been shown to be more active in comparison with FOS. The character of their influence on lactobacilli is discussed and the conclusion is made that the restoration and maintenance of eubiosis is greatly determined by the pool of metabolites excreted by the bacteria.     

Apparent minimal size required for cell division in Escherichia coli.

The experiments described in this report were designed to find out whether there is a minimal size threshold for cell division or for DNA replication in Escherichia coli. Cells with decreasing size (or mass) were obtained by successive amino acid starvations. Following two starvations, the cells were at least 30% smaller than unstarved newborn cells. The results suggest that this size is below a minimal size threshold for cell division but not for initiation of DNA replication.     

Lipid synthesis during the Escherichia coli cell cycle.

Lipid synthesis was examined in Escherichia coli cells at different stage of cell division. Exponentially growing cells were pulse-labeled with appropriate isotopes for 0.1 generation time, inactivated, and separated by size on a sucrose gradient. An abrupt increase in the rate of lipid synthesis occurred which was coincident with the initiation of cross walls. In contrast, the rate of protein synthesis during this same interval remained constant, resulting in an increased lipid/protein ratio in dividing cells. No changes in the composition of phospholipid head groups, fatty acids, or phospholipid molecular species were observed in cells at different stages of division. The observed increase in the rate of lipid synthesis may reflect a means by which the activities of membrane-associated enzymes are modulated during cross wall formation.     

Flow cytometric narrow-angle light scatter and cell size during starvation of Escherichia coli in artificial sea water

Flow cytometry was used to study starvation of Escherichia coli in artificial sea water. Flow cytometric narrow-angle light scatter was compared and assessed in relation to the cell sizes obtained by scanning electron microscopy at low temperature, and by image analysis. A correlation between narrow-angle light scatter and cell size was not observed, although an acceptable correlation (γ= -0.845) between narrow-angle light scatter and the starvation period was observed. On the other hand, the distribution of narrow-angle light scatter at any given moment of culture is asymmetric and may be associated with the cell size distribution at the specific moment of starvation.     

Effect of Actoflor preparations on survival of Escherichia coli M-17 and Salmonella enteritidis under starvation conditions in mixed cultures

The viability of E. coli M-17 and S. enteritidis under starvation conditions in mono- and mixed cultures was studied. E. coli M-17 showed greater capacity for survival in mixed cultures than in monocultures, while for S. enteritidis the contrary was true. Preparations "Actoflor" enhanced the antagonistic activity of E. coli M-17, ensuring its absolute selective advantage under starvation conditions in mixed cultures. The role of E. coli M-17 low-molecular exometabolites is discussed; they are probably an important factor in the antagonistic activity of these microorganisms.